1. Field of the Invention
The present invention relates to agene, especially a cloned gene coding for a interleukin-2polypeptide, recombinant DNA carrying the gene, a living cell line possessing the recombinant DNA and a method for producing interleukin-2 using the cell line.
2. Brief Description of the Prior Art
Interleukin 2 (hereinafter referred to as "IL-2"), formerly referred to as T cell growth factor, is a soluble protein (generally known as "lymphokine"), and is produced froin T cells activated with a lectin or an antigen (Morgan. D. A., et al., Science, 193, 1007-1008 (1976), Gillis, S. et al., J. Immunol., 120, 2027-2033 (1978). Interleukin 2 (IL-2) is capable of modulating lymphocyte reactivity and promoting the in vitro long-term culture of antigen specific effector T-lymphocytes (Gillis. S. et al., Nature 268, 154-156 (1977)). IL-2 is also known to manifest other relevant biological activities such as enhancement of thymocyte mitogenesis (Chen, B. M. et al., Cell. Immunol., 22, 211-224, (1977), Shaw, J. et al., J. Immunol. 120, 1967-1973, (1978)), induction of cytotoxic T cell reactivity (Wagner, H. et al., Nature, 284, 278-280, (1980)) and anti-SRBC plaque forming cell responses (Gillis, S. et al., J. Exp. Med., 149, 1960-1968, (1979)) in cultures of nude mouse spleen cells. Accordingly, this lymphocyte regulatory substance is useful in potentiating humoral and cellular immune responses and in restoring immune deficient state to a normal humoral and cellular immune state. These identified immunological activities of IL-2 strongly indicate that IL-2 is useful for medical immunotherapy against immunological disorders including neoplastic diseases, bacterial or viral infections, immune deficient diseases, autoimmune diseases etc.(Papermaster, B. et al., Adv. Immunopharm., 507, (1980)). Like inteferons, IL-2 has been shown to augment natural killer cell activity, suggesting a potential use in the treatment of neoplastic diseases. Furthermore, IL-2 enables the maintenance of cultures of functional monoclonal T cells and hence appears to play a key role in the studying of the molecular nature of T cell differentiation, and of the mechanism of differentiated T cell functions as well as the mechanism of T cell antigen receptors. It is also useful for producing, by long term culturing of monoclonal T cell, many other T cell derived lymphokines, which are useful in a wide range of fields. In addition, IL-2 production and the response of lymphocytes to IL-2 can be important parameters of immunological functions which are useful in the clinical diagnosis of aberrant immunity.
IL-2 has been produced in the prior art by stimulating mouse, rat or human lymphocytes with a mitogen (Gillis. S. et al., Nature, 268, 154-156, 1977, Farrar, J. et al., J. Immunol., 121, 1353-1360,(1978). Gillis, S. et al., J. Immunol., 120, 2027-2033, 1978,)). By stimulating human peripheral blood mononuclear lymphocytes with a mitogen (Gillis. S. et al., J. Immunol., 124, 1954-1962, (1980)). Gillis et al. reported the preparation of murine IL-2 from murine T cell lymphoma cell line (Gillis. S. et al, J. Immunol., 125, 2570-2578 (1980)) and the preparation of human IL-2 from a human leukemia cell line (Gillis, S. et al., J. Exp. Med., 152, 1709-1719, (1980)).
The above noted articles by Gillis et. al. discuss the method of producing human IL-2 from mitogen-stimulated human T cell leukemia cell line by cell culture methods. However, such a technique results in undesirably low concentrations of human IL-2, and necessiates complex purification procedures to obtain even small amounts of IL-2 from a huge volumes of culture media. Moreover, since the human T cell leukemia cell lines produce trace amounts of many other biologically active substances which are analogous to human IL-2, significant difficulties are encountered in isolating IL-2 from these other immunologically active molecules, or in isolating IL-2 from the occasionally present toxic lectins.
As an alternative approach it would seem to be desirable to use recombinant DNA (DNA is an abbreviation for deoxyribo-nucleic acid) techniques as are used in the production of other biologically active human proteins, such as interferons, (Gray, P. W. et al., Nature, 295, 503-508, (1981), Nagata, S., et. al., Nature, 284, 316-320, (1980), Taniguchi, T. et. al., Gene, 10, 11-15, (1980)) to produce IL-2. However to date, attempts at the production of IL-2, by recombinant DNA techniques have not been successful. For instance, it was reported in "NIKKEI BIOTECHNOLOGY (Japan), No. 19, Jul. 5, 1982 that attempts to construct IL-2-producing organisms by recombinant DNAwere unsuccessful, probably due to the fact that the gene coding for IL-2 polypeptide had not yet been cloned.
A need therefore, continues to exist for a cloned gene, coded for interleukin-2, and for DNA produced recombinantly which carries the gene. A need also continues to exist for a living cell line which possesses the recombinantly produced DNA, and for a method of producing interleukin-2 using the cell line.